Metformin potentiates DSF/Cu-loaded pluronic nanoparticles to improve therapeutic efficacy in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype characterized by poor prognosis and resistance to conventional therapies, underscoring the need for novel therapeutic strategies. Drug repurposing has emerged as a promising approach, with disulfiram (DSF) and metformin demonstrating anticancer potential. In addition, nanoparticle (NP)-based drug delivery systems can enhance therapeutic efficacy and tumor selectivity. This study investigates the antitumor effects of a combination therapy using DSF/Cu complex–loaded pluronic-based nanoparticles and metformin in TNBC cells. Pluronic-based nanoparticles were synthesized using the thin-film hydration method, and hyaluronic acid (HA) was conjugated to pluronic, as confirmed by H-NMR spectroscopy. Drug–nanoparticle interactions were characterized using FTIR analysis. Drug loading efficiency, release profiles, and cellular uptake were evaluated under acidic and physiological conditions. Cytotoxic effects of free drugs and NP formulations were assessed using the MTT assay, while apoptosis was analyzed by flow cytometry. The expression levels of STAT3, CD24, CD44 and ALDH1 were quantified by real-time PCR. Successful conjugation of HA to pluronic was confirmed by H-NMR, and FTIR analysis verified physical interactions between the drugs and pluronic micelles. Encapsulation efficiencies for DSF and DSF/Cu were 89.5% and 73%, respectively. DSF/Cu-loaded nanoparticles exhibited a faster drug release under acidic conditions compared to physiological pH. NP formulations significantly enhanced cellular uptake and demonstrated a synergistic-like potentiating effect, wherein sub-cytotoxic concentrations of metformin markedly enhanced the anticancer efficacy of DSF/Cu-loaded nanoparticles. Apoptosis assays revealed increased apoptotic cell death with minimal necrosis. Furthermore, real-time PCR analysis showed significant downregulation of STAT3 and stemness-associated gene expression compared to untreated controls. Pluronic-based nanoparticles efficiently encapsulate DSF/Cu and enhance its anticancer activity. The combination of metformin with DSF/Cu-loaded nanoparticles significantly inhibits TNBC cell growth and reduces stemness properties. This low-toxicity, metabolism-assisted combination strategy shows strong potential for TNBC treatment and warrants further in vivo investigation.